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The Center offers instrumentation for injection of fluorescent dyes, proteins, and small molecules into living cells with subsequent real-time microscopic observation for fluorescence or FRET. The set up includes an Eppendorf 5171 and 5246 Microinjection and Micromanipulation system on a Zeiss Axiovert 135 TV microscope with phase contrast optics to follow microinjection. This system is equipped with an Attofluor Dual Intensified ICCD camera module and third CCD camera for observation during microinjection. Eppendorf Femtotips are available for microinjection or glass pipettes can be produced on site with a Sutter P97 Horizontal Pipette puller.
This system can be used for a wide variety of experimental protocols, which can be set up and stored in the software. While microinjection of substances into cells is not a high throughput technique, it can often be the fastest way to observe the intracellular actions of substances not readily taken up by cells.
Once dyes are injected, the two sensitive high resolution cameras can simultaneously detect two wavelengths of emission allowing for accurate dual-emission fluorescence imaging and FRET. A filter changer can rapidly switch (in a manual or automated mode) the excitation wavelengths in an experimental sequence. Images are recorded and the fluorescence intensity of Regions of Interest (ROIs) can be followed over time as directed by the software. |
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